Mechanosensitivity of the detrusor is defined as the ability of smooth muscle cells to generate mechanical activity independent of external stimuli. During bladder filling, there is usually no parasympathetic outflow from the spinal cord, however, the bladder develops tone and also exhibits non- synchronized local contractions and relaxations. Pathological changes in mechanosensory mechanisms may lead to the development of detrusor overactivity (DO) which is a co-symptom of several dysfunctions of the lower urinary tract including overactive bladder, obstructed bladder, urinary incontinence, and bladder pain syndrome. The central hypothesis of this proposal is that impaired mechanosensation and mechanotransduction in bladder smooth muscle cells (BSMC) results in the abnormal response of the human bladder to physiological stretch and detrusor overactivity due to the changes in mechano-gated two-pore domain (K2p, KCNK) K+ channels. Animal data and our preliminary results from the human detrusor suggest that stretch-activated two-pore domain (K2p, KCNK) K+ channels play a critical role in bladder mechanosensitivity. We also established that TREK-1 channel is a predominantly expressed member of stretch-activated K2p channels in the human detrusor. Specific Aim 1 will identify differences in the level of expression and function of stretch-activated K+ (K2p) channels in human bladder smooth muscle cells in patients without and with detrusor overactivity. Specific Aim 2 will evaluate interactions between TREK-1 and cytoskeleton (actin microfilaments, caveolin, membrane lipids) in human BSMC and compare the role of TREK-1 modulating proteins in normal vs. overactive human detrusor. Specific Aim 3 will test the efficacy of gene therapies (gene silencing by siRNA and gene activation by saRNA) in modulating the expression and function of TREK-1 in human BSMC in order to effectively regulate an abnormal response of the overactive detrusor to stretch during bladder filling. The overall objective of this proposal is to investigat cellular and molecular mechanisms of abberant mechanosensitivity and altered response of the human detrusor to physiological stretch associated with increased myogenic tone and spontaneous non-voiding contractions in patients with DO. Proposed studies will clarify the cellular mechanisms of mechanosensitivity and mechanotransduction in the human bladder and in patients with idiopathic DO, and provide new information for the development of new pharmacological interventions and innovative strategies for the treatment of urinary bladder dysfunctions.